Neisseria gonorrhoeae (Ng) is the causative agent of the sexually transmitted infection (STI), gonorrhea. Worldwide, more than 106 million new cases occur annually. Ng causes natural infection exclusively in humans and is a serious public health problem and has dire implications for the reproductive health of women. Treatment of this infection has become resistant to almost every antibiotic that has been used; development of new therapies has lagged behind and Ng has now achieved superbug status. The emergence and spread of ceftriaxone-resistant strains, in particular, has heralded an era of untreatable gonorrhea and represents a global public health problem. There is an urgent need to develop novel vaccines against this pathogen. Ab elicited by immunization of mice with a peptide mimic ('mimitope') of a lipooligosaccharide structure recognized by mAb 2C7 (the 2C7 oligosaccharide epitope) configured as a multi-antigen peptide (2C7-MAP) mediates direct complement (C) dependent killing and opsophagocytosis by polymorphonuclear leukocytes (PMNs) in vitro and hastens clearance of Ng in the mouse vaginal colonization model. In Specific Aim 1, 2C7-MAP that has been synthesized to a purity of >95% will be tested for immunogenicity using adjuvants currently in clinical use. The immunogenicity following routes of administration employed in humans will also be studied. Gonorrhea is a host-restricted infection. Several factors, including human-specific complement (C) evasion may contribute to the tropism of Ng for humans. Ng binds to C inhibitors, factor H (fH) and C4BP in a human-specific manner; binding of these inhibitors can thwart the efficacy of vaccine Ab. In Specific Aim 2, we will use novel human fH and C4BP transgenic (Tg) mice that we have developed to evaluate the efficacy of immune Ab elicited by our vaccine candidate in the face of the possible obstacles posed by human fH and C4BP, as would be encountered in humans. Because binding of fH and C4BP to gonococci is human-specific we believe that vaccine efficacy in fH/C4BP Tg mice will more faithfully simulate protection in humans. PMN influx will also be quantified. Clinical gonococcal infection is characterized by an influx of PMNs into the genital tract, which in concert with Ab and complement may facilitate clearance of infection. In Specific Aim 3, we will elucidate the roles of PMNs, complement and complement receptors in vaccine efficacy. Experiments will be performed in BALB/c mice from which PMNs are depleted, or C3 (opsonin) and C5a receptor (receptor for the key complement chemotaxin, C5a) are deleted. Completion of the proposed studies will represent an important step forward in the development of novel vaccines against gonorrhea.